Chemically Synthesized SDF-1α Analogue, N33A, Is a Potent Chemotactic Agent for CXCR4/Fusin/LESTR-expressing Human Leukocytes

Abstract

Stromal cell-derived factor (SDF) 1 is a potent chemoattractant for leukocytes through activation of the receptor CXCR4/Fusin/LESTR, which is a fusion co-factor for the entry of T lymphocytotropic human immunodeficiency virus type 1 (HIV-1). This CXCR4-mediated HIV-1 fusion can be inhibited by SDF-1. Because of its importance in the study of immunity and AIDS, large scale production of SDF-1 is desirable. In addition to recombinant technology, chemical synthesis provides means by which biologically active proteins can be produced not only in large quantity but also with a variety of designed modifications. In this study, we investigated the binding and function of an SDF-1␣ analogue, N33A, synthesized by a newly developed native chemical ligation approach. Radioiodinated N33A showed high affinity binding to human monocytes, T lymphocytes, as well as neutrophils, and competed equally well with native recombinant SDF-1␣ for binding sites on leukocytes. N33A also showed equally potent chemoattractant activity as native recombinant SDF-1␣ for human leukocytes. Further study with CXCR4/Fusin/LESTR transfected HEK 293 cells showed that N33A binds and induces directional migration of these cells in vitro. These results demonstrate that the chemically synthesized SDF-1␣ analogue, N33A, which can be produced rapidly in large quantity, possesses the same capacity as native SDF-1␣ to activate CXCR4-expressing cells and will provide a valuable agent for research on the host immune response and AIDS.

The murine chemokine receptor CXCR4 is tightly regulated during T cell development and activation

“…Chemotaxis toward SDF-1␣ has been reported for various leukocyte populations, including hematopoietic progenitor cells, T cells, B cells, monocytes, and neutrophils [7,[18][19][20][21]. Among various organs, SDF-1 was found to be transcribed in the thymus.…”

“…5]. Expression of human CXCR4, one of those receptors, has been observed on different cells of the immune system including B cells, monocytes, macrophages, Langerhans cells, neutrophils, and T cells [6][7][8][9]. In the thymus, expression of CXCR4 was found on all stages of T cell development including doublepositive (CD4 ϩ CD8 ϩ ) and single-positive (CD4 ϩ CD8 Ϫ ; CD4 Ϫ CD8 ϩ ) T cells [8].…”

Post‐translational and cell type‐specific regulation of CXCR4 expression by cytokines

“…While some authors were unable to detect expression of CXCR4 [6,7], others have described binding of SDF-1 § , SDF-1 § -induced Ca 2+ flux and migration [8][9][10]. We have detected a clear expression of CXCR4 on neutrophils from the synovial fluid of patients with different inflammatory joint diseases, whereas CXCR4 was hardly detectable in the peripheral blood of patients or normal blood donors, suggesting that CXCR4 on neutrophils could be functionally relevant in the setting of inflammation [11].…”

Receptor reserve analysis of the human CCR3 receptor in eosinophils and CCR3-transfected cells

“…Guinea pig eotaxin was made synthetically by Gryphon Sciences (South San Francisco, CA), as has been done with other chemokines [25]. 125 I-Eotaxin was obtained from New England Nuclear (Boston, MA).…”